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Membrane Stripping and Re-probing Protocol - Western Blotting

 

Membrane Stripping and Reprobing Protocol for Western Blotting

 

    Stipping and re-probing blots

    • add 280 μl of 2-beta-mercaptoethanol per 40 mL of stripping buffer (to a final concentration of 100 mM)
      • Wash membrane in TBST (1 X 5 min)
      • Incubate membrane in stripping buffer for 30 min at 50-60°C (use a heating oven for this - and keep the oven door closed due to smell)
      • Wash in TBST (2 X 5 min)
      • Block membrane for 1 h in 5% blocking milk
      • Follow normal protocol for developing western blots.
      • Membrane Stripping Buffer Recipe:
      • Stripping buffer: 62.5 mM Tris-HCl (pH 6.8), 2% SDS. (add 280 ul of 2-beta merceptoethanol or a final concentration of 0.1 M 2-Mecaptoethanol).
      • See our Western Blotting Protocol


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